Abstract

Bio-inspiration for novel adhesive development has drawn increasing interest in recent years with the discovery of the nanoscale morphology of the gecko footpad and mussel adhesive proteins. Similar to these animal systems, it was discovered that English ivy (Hedera helix L.) secretes a high strength adhesive containing uniform nanoparticles. Recent studies have demonstrated that the ivy nanoparticles not only contribute to the high strength of this adhesive, but also have ultraviolet (UV) protective abilities, making them ideal for sunscreen and cosmetic fillers, and may be used as nanocarriers for drug delivery. To make these applications a reality, the chemical nature of the ivy nanoparticles must be elucidated. In the current work, a method was developed to harvest bulk ivy nanoparticles from an adventitious root culture system, and the chemical composition of the nanoparticles was analysed. UV/visible spectroscopy, inductively coupled plasma mass spectrometry, Fourier transform infrared spectroscopy and electrophoresis were used in this study to identify the chemical nature of the ivy nanoparticles. Based on this analysis, we conclude that the ivy nanoparticles are proteinaceous.

AFM and SEM images of ivy nanoparticles. (a) AFM scan of dense ivy nanoparticles secreted directly from an adventitious root. (b) AFM scan of dense ivy nanoparticles isolated using the procedure developed in this study. (c) Small cluster of ivy nanoparticles imaged by AFM after dilution from the concentrated sample collected from the column. The inset of (c) shows an SEM image of a single ivy nanoparticle prepared the same as the diluted AFM sample. Note that the size of an individual nanoparticle is slightly smaller by AFM; however, artefacts related to tip–particle interactions can greatly affect size measurements using AFM.

DLS and zeta potential analysis of the isolated ivy nanoparticles. (a) DLS of the nanoparticles collected from three separate isolations showed a similar distribution, with a mean diameter of 95.69 ± 5.56 nm. (b) The zeta potential of the ivy nanoparticles was found to be −35.3 mV, indicating that the ivy nanoparticles did not form a stable solution in ultrapure water. (Online version in colour.)

(a,b) Peaks observed from UV detector of the ivy extract. A prominent peak was observed in both wavelengths (highlighted) during the 10–11 min fraction. This fraction corresponded to the presence of nanoparticles, as indicated by AFM. Peaks with lower intensity were imaged but were found not to contain any nanoparticles. (Online version in colour.)

(a) UV/vis spectra of the ivy nanoparticle fraction collected directly from the HPLC column. Note the wide absorbance from 200 to 350 nm, before dropping off in the visible region. (b) A plot of absorbance versus concentration at 283 nm clearly shows the direct effect of the nanoparticle concentration on the absorbance. (Online version in colour.)

Diagrammatic representation of the results from the ICP-MS and elemental analysis. As indicated, the C : N ratio was approximately 10 : 1, indicating that the nanoparticles were composed of biomolecules. Additionally, ICP-MS revealed that all metals in the ivy nanoparticle fraction were less than 37 ppm, confirming that the ivy nanoparticles are organic.

FTIR spectrum of the ivy nanoparticles. The FTIR spectrum for the ivy nanoparticles was compared with reference spectra for chitosan (a representative polysaccharide) and BSA (a representative protein). All three samples had a band at 1653 cm−1, indicating vibration around the CO–NH bond, and around 2928–2932 cm−1 indicating C–H vibration. In addition, the ivy sample shared a peak at 1071–1076 cm−1 with the chitosan sample, indicating vibration of a CO–C bond, typical of sugars. This band was not present in the BSA sample. Similarly, the BSA sample had a strong peak at 1518 cm−1, representing the amide II band, whereas the ivy nanoparticles had a weak band at 1539 cm−1, indicating a weak amide II band, and the chitosan sample had no peak in this region. The FTIR spectra from top to bottom are: raw ivy nanoparticles, chitosan and BSA. (Online version in colour.)

Results from SDS-PAGE of ivy nanoparticles. (a) Results of the silver stain demonstrating the staining of the protein ladder, and all ivy nanoparticle samples. (b) Results from glycoprotein stain showing positive staining for the high MW nanoparticle band. Samples 1, 2 and 3 represent nanoparticles isolated from three separate trials. Note the lack of staining of the non-glycosylated proteins from the standard ladder.